Recent data suggest that the local circuitry of the developing brain may be molded by an activity-dependent molecular mechanisms that capitalizes on the unique ligand and voltage-gated properties of the NMDA subtype of glutamate receptor. Our studies use the retinotectal pathway of amphibian larvae as a model system for studying the mechanisms of activity dependent competition within the visual system. Our recent evidence indicates that chronic disruption of NMDA receptor function blocks the fine-tuning of the retinotopic map in normal tecta and causes desegregation of ocular dominance stripes in doubly innervated tecta. The data suggest that NMDA receptor activation increases the lifetime of retinal synapses carrying correlated activity and simultaneously decreases the probability of new sprouts from retinal terminals in the vicinity of the synchronized synapses. However, without EM analysis we cannot determine whether synaptic changes are actually involved in the light microscopic alterations we observe with double innervation and drug treatment. We propose using a newly developed sampling technique for analyzing the distribution of synapses within individual HRP labeled retinal ganglion cell arbors to determine whether changes in the distribution, size, morphology, or the amount of synaptic contact supported by individual ganglion cell terminals accompany the pronounced NMDA related changes in retinal terminal morphology that we observe. We will also use this technique, EM analysis of retinotectal contacts of known lifetime in cocultures of retinal explants and tectal cells, and EM immunocytochemistry using Synapsin I and stereological sampling procedures to determine if distinct morphological correlates of new synapses, synapses in the process of retraction and stabilized or "long-lifetime" synapses exist. Finally we will determine the types of synaptic relationships that developing retinal afferents participate in using MAP 2 antibody to identify dendrites, our own monoclonal marker for axons and HRP label of retinal synapses, and we will attempt to use a new combined physiological and EM technique to identify the processes in the retinotectal neuropil with the highest concentrations of NMDA receptors by virtue of their ability to concentrate Ca++ ions. We believe that this work represents a crucial step in unraveling the cascade of synaptic and biochemical interactions that mediate activity- dependent competition in all vertebrate central visual pathways. Understanding these interactions will ultimately be crucial to the development of effective clinical procedures for ameliorating the destructive effects of abnormal early experience or of countering the effects of lesions in the human visual pathway.